1. Field of the Invention
The present invention relates to magnetic nanoparticles, and particularly to magnetic nanoparticles for protein immobilization.
2. Description of the Related Art
Magnetic particles are being increasingly used as carriers for binding proteins, enzymes, and drugs. Such immobilization procedures for proteins and other biologically active compounds have a major impact in different areas of biomedicine and biotechnology. For example, nano magnetic beads or particles are used in various applications in medical, diagnostic and industrial applications. Coupling of proteinaceous sources at the surface of magnetic beads has been achieved by several methods using surface activating material and surface functionalized magnetic particles. Surface activating chemicals, such as ethyl (dimethylaminopropyl) carbodiimide (EDC) react with surface carboxylated magnetic beads and the resulting chemical group is highly reactive with the amino groups in proteins. However, the main disadvantage of this method is the high cost and instability of the EDC compound, which therefore limits the adoption of this technology on a commercial scale.
Coating of magnetic beads provides an alternative avenue for coupling with proteins by providing a wide variety of functional groups (aldehyde, epoxy, etc.) that can be used directly in protein coupling. Chitosan compound was used previously for coating magnetic beads, as a heavy metal ion capture, in water treatment in order to couple various proteins for diagnostic and medical usages. Chitosan β-(1-4)-linked D-glucosamine is the second most abundant material after cellulose and has been widely used in various applications as a biodegradable and environmental friendly material. Chitosan-coated magnetic beads offer a low cost option for large scale production of enzyme-coupled magnetic nanoparticles in various applications. Typically, however, preparation of chitosan-coated magnetic beads often includes activating the magnetic bead's surface by EDC and cross-linking it by glutaraldehyde. While this protocol has been used for large-scale production of magnetic beads, use of EDC renders the process costly.
Reverse phase emulsion is a preparation method for magnetic nanoparticles which facilitates control of the size (diameter) of the magnetic particles. Glutaraldehyde is added directly after the formation of the emulsion. The nanoparticles are then be purified for surface activation and enzyme coupling. Some researchers have activated the surface of chitosan-coated beads by incubating it with a low amount of glutaraldehyde and then added enzyme after washing. The disadvantage of this method, however, is the intra cross-linking between amino groups across magnetic beads after activation. This type of cross-linking can yield clotted beads which can affect the homogeneity in the diameter of the beads.
Thus, a method for preparing chitosan-coated magnetic nanoparticles for protein immobilization solving the aforementioned problems is desired.